Abuse of prescribed and other sedative drugs (e.g., benzodiazepines, barbiturates and ethanol) is among the top five health problems in the U.S. and is one of the most highly heritable addictive disorders. A host of biological (genetic) and environmental factors interact throughout the addictive process to influence drug use/abuse. Physiological dependence and associated withdrawal episodes are thought to constitute a motivational force that perpetuates sedative drug use/abuse and contributes to relapse. In humans, the identification of genes that influence drug dependence and withdrawal has been limited. Thus, the use of preclinical (animal) models that closely approximate the clinical situation is essential to elucidate the gene networks involved. We used these methods to identify two genes involved in sedative withdrawal in mice. The human homologs are now being studied in human populations by NIH intramural and other scientists. Quantitative trait loci (QTLs) are chromosome sites containing genes that influence a complex trait such as predisposition to drug withdrawal and consumption. Previously, we identified QTLs on chromosomes 1 and 4 with a large effect on sedative withdrawal in mice. During the current funding period, we fine-mapped these QTLs to 0.44 and 1.8 Mb intervals (syntenic with human 1q23.2 and 9p24-p22.3) and identified genes that code for MPDZ (the multi-PDZ domain protein) and GIRK3 (G-protein activated inwardly rectifying potassium channel subunit) as quantitative trait genes for drug withdrawal. Analyses of GIRK3 and MPDZ genetic models suggest that GIRK3 and MPDZ may also affect other addiction relevant phenotypes including sedative and cocaine self-administration. A key hypothesis of this proposal is that GIRK3 and MPDZ affect withdrawal from and self-administration (consumption) of multiple drugs of abuse. A second hypothesis is that the molecular network(s) associated with GIRK3 and MPDZ effects on behavior may interact at a mechanistic level. The aims may be summarized: 1) To elucidate the roles of MPDZ and GIRK3 in drug withdrawal and self- administration using novel MPDZ and GIRK3 genetic models. 2) To elucidate brain regions involved in GIRK3 and MPDZ effects on behavior using chemical lesions as primary procedures. 3) To test the role of GABAB and 5-HT2C receptors in GIRK3 and/or MPDZ actions with central administration of selective agonist/antagonists to GIRK3 and MPDZ genetic models and wildtype littermates. 4) To elucidate the MPDZ and GIRK3 network signalosomes using weighted gene co-expression network analysis (WGCNA). Gene expression will be assessed using the Illumina array (e.g., beginning with the central nucleus of the amygdala) utilizing novel MPDZ and GIRK3 genetic models, using withdrawn and control mice. In addition to identifying differentially expressed genes, innovative de novo network construction will allow us to elucidate the underlying regulatory structure at a high level of integration. This can direct further mechanistic studies and elucidate the important network(s), within which many biomarkers and drugable targets may exist to direct translational studies. PUBLIC HEALTH RELEVANCE: The proposed studies address genetic mechanisms involved in drug physiological dependence and associated withdrawal, and their broader role in addiction-relevant behaviors including self-administration. Elucidating the genetic bases of these mechanisms will provide a significant boost not only in understanding the underlying pathology but also in the development of new treatment strategies.